With rapid development of molecular biology and constant improvement of genetic cloning technology, studies on genetic engineering for plants and microbes are being developed in depth and breadth, and researches on resistance genes have been transferred from tolerance to biotic stresses (e.g. disease, pest) to tolerance to abiotic stresses, such as drought, acid-alkaline, saline-alkaline and heat.
Because of the increased CO2 emission, greenhouse effect on the earth is growing worse and leading to global warming. It is estimated that the global average temperature will increase by 1.4-5.8° in the next 100 years. Global warming gradually deteriorates the agricultural ecological environment. It is predicted that climate warming may lead to 17% of crop yield reduction. One research from IRRI (International Rice Research Institute) proved that during 1998-2003, the crop yield was decreased by 10% with the temperature elevated by 1°. In China, experts believe that by 2050, the nationwide average temperature will increase by 2.2°. Plants growing under natural conditions are all affected by the elevated temperature and grow more slowly. Some major crops, such as rice and corn, are especially easy to be influenced by hot weather during heading and filling stage, and result in crop yield reduction. On the other hand, according to FAO (Food and Agriculture Organization of the United Nations), the world population will exceed 10 billions by 2050. With further increased world population, there will be more and more pressure on agriculture, and worldwide food shortage will be a long-lasting problem. Being affected by global warming, lots of herbaceous plants will grow more slowly and even die, thus breaking ecosystem balance. Therefore, scientists all over the world are taking great efforts in searching for heat-tolerance relevant plant genes. So far from now, only a few heatshock protein genes and transcription factors thereof are found to be relevant with heat tolerance, while none of a single gene was reported to be capable of increasing heat tolerance of bacteria and plants.
Nowadays, there are 1 billion hm2 of saline-alkaline land, which is about 10% of global arable areas, in more than 100 nations in the world. China alone has 99.13 millions hm2 of saline-alkaline land, mainly at arid and semiarid regions of the north, northwest and northeast of China. There are more than 3.70 millions of hm2 of saline-alkaline land at Songnenpingyuan at the western part of northeast China, which is one of the three major centralized areas of soda saline-alkaline land. Meanwhile, the areas of secondary salinization land are increasing rapidly due to the industrial pollution, the irrational irrigation and the ill use of chemical fertilizers. Saline-alkaline land affects the vegetation growth by reducing or even terminating the crop output, and it also indirectly deteriorates the ecological environment and corrodes engineering installations, which leads to 2.511 billion yuan of losses every year. Therefore, it is one of the problems that demand urgent solution in sustainable development of agriculture to reduce damages of soil salination to crops and make full use of the limited land resource. Besides of the comprehensive treatment by traditional physical, chemical and biological ways, etc., it will be one of the most cost-effective methods for enhancing tolerance of plants to stresses by genetic engineering with the up-to-date molecular biology method.
Saline-alkaline soil is the soil that contains too many salts of NaCl, Na2SO4, Na2CO3 and NaHCO3. Damages of saline-alkaline soil towards plants mainly include complexed damages from stresses of salinity, high-pH and the interaction thereof. Damages from saline-alkaline stress are mainly represented in three ways: first, the massive accumulation of metal ions (mainly Na) in cytoplasm, which breaks the ionic balance and inhibits physiological and biochemical metabolic processes in cells, thus weakening the photosynthesis ability of plants and finally killing them with carbon starvation; second, the high osmotic circumstance of saline-alkaline soil, which may stop plant root systems from absorbing water, thus causing plants to die from “drought”; third, the relatively high pH value of saline-alkaline soil, which disturbs the acid-base balance between plants and the external environment, thus disrupting the membrane structure of cells and killing plants with exosmosis of cell content. Therefore, plants under saline-alkaline stresses need, on one hand, to reduce ion accumulation in cytoplasm; on the other hand, to generate from accumulation process some special products, such as proteins, amino acids and sugars, to increase osmosis of the cell, thus preventing water losses and stabilizing structures of plasma membrane and enzymes.
Being widespread, saline-alkaline land is becoming a new hotspot of research. Studies now are mainly focused on how plants on saline-alkaline land respond to pH stress, whilst there is only preliminary exploration towards physiological characterization and gene expression. The major objects of the study are certain kinds of saline-alkaline tolerant plants, such as weeping bulrush (Puccinellia tenuiflora), chinensis (Leymus chinensis), sunflower (Helianthus annuus) and nitrebush (Nitraria schoberi). However, studies of plant response to high pH stress at the molecular level are processing slowly. There are demands in the art for the development of backup genes that can enhance the tolerance of plants to saline-alkaline stress, as well as methods for enhancing the tolerance of plants to saline-alkaline stress by genetic engineering techniques.
The environmental hydrogen potential is normally presented by the negative logarithm of hydrogen ion concentration, i.e., pH value. The environmental pH greatly affects the vital movement of microbes on that: pH variation changes the electric charge on the surface of microbes, thus affecting microbe absorption towards nutrients; pH can affect the ionotropy of organic compounds in culture medium besides of direct influences to microbe cells, thus affecting microbes indirectly, since most non-ionic compounds penetrate into cells more easily than ionic ones; only with optimum pHs can maximum activity of enzymes be achieved, and those unsuitable pHs decrease enzyme activities and therefore affect the biochemical processes in microbe cells; and, pHs of too high or too low will both reduce the tolerance of microbes to heat.
With the growth of microbes in substrates, the hydrogen ion concentration of substrates will be changed with metabolism. As environmental pH changes, growth of microbes is retarded, and pHs beyond the maximum or minimum of tolerance will lead to death of microbes. With rapid development of molecular biology and constant improvement of genetic cloning technology, microbes with resistance can be cultivated through engineering studies that are being developed in depth and breadth, the key point of which is to find tolerance genes of microbes to saline-alkaline stresses.
Water resource shortage is now a global problem that restricts the development of agriculture. According to statistics, there are about 43% of arable lands that are under stresses of drought and semi-drought. The drought stress not only severely affects the growth of crops and reduces the yield, but also limits the promotion of improved crop strains. Therefore, it is one of the hot issues to enhance the tolerance of crops to drought in modern agriculture studies.
Studies on drought tolerance of plants relate to many fields such as plant morphology, physiology and biochemistry as well as molecular biology. It has been paid close attention to studies on drought tolerance with the following aspects, namely, structure changes of plant root systems and leaf blades under drought conditions; relationship between abscisic acid (ABA) and stomatal closure; relationship between drought tolerance of plants and osmoregulation substances of small molecule compounds, such as mannitol, proline, betaine, trehalose, fructosan, inositol, polyamine, etc.; and effects of aquaporin, reactive oxygen removal and late embryogenesis abundant protein on drought tolerance of plants.
With the development of molecular biology research, some important drought tolerant genes are discovered and cloned one after another, and drought tolerant transgenic plants of tobacco and rice are obtained. Transgenic rice lines with drought tolerance have been successfully cultivated, which brought about broad utilization prospects on studies of drought tolerant genes of other plants. Now, there are mainly two strategies for cultivating drought tolerant species by genetic engineering techniques. One is to enhance the synthesis capacity of permeable metabolites of plants, which can therefore synthesize under water stresses more osmoregulation substances (e.g. mannitol, betaine, trehalose, etc.) to improve the osmoregulation, thus enhancing the drought tolerance of plants. The other is to enhance the ability of plants of clearing active oxygen radicals with over expression of certain enzymes (e.g. SOD, POD, CAT, etc.) under water stresses, thus getting rid of harmful active oxygen radicals effectively and enhancing the drought tolerance of plants. With osmoregulation as a main mechanism for the drought tolerance of plants, improvement for synthesis of proline and betaine has recently been achieved with plant genetic engineering method, and promising progresses have been made in the cultivation of drought tolerant transgenic plants largely based on osmoregulations.
Proline is an amino acid of great solubility. With dipolarity, proline connects proteins with its hydrophobic end and water molecules with hydrophilic end, thus binding more water molecules for proteins and increasing the solubility thereof to involve more soluble proteins in osmoregulations. Meanwhile, the improvement of bound water content may prevent or decrease protein denaturations caused by dehydration of cells. Therefore, the improvement of synthetic ability for proline may enhance the drought tolerance of plants, and some successful reports have been made in this respect.
All in all, it is recently a hotspot of improving plants with genetic engineering techniques, and it is one choice to enhance the plant tolerance to abiotic stresses and cultivate plant lines of resistance by genetic engineering techniques. However, there are seldom reports of a single gene that could comprehensively enhance various tolerances of plants and microbes to abiotic stresses.